i. Biological Tissue Grafts and Prostheses
Collagen, and to a lesser extent elastin, are the major connective tissue proteins which make-up the connective tissue framework of most biological tissues. The relative pliability or rigidity of each biological tissue is largely determined by the relative amounts of collagen and elastin present in the tissue and/or by the physical configuration and conformation (e.g., structural lattice) formed by the connective tissue proteins.
The prior art has included numerous surgically-implantable bioprosthetic grafts and prostheses (referred to herebelow collectively as "bioprostheses") which are formed wholly or partially of chemically fixed (i.e., tanned) biological tissue. The chemical fixation of these biological tissues is typically accomplished by contacting the tissue with one or more chemicals which will crosslink collagen and elastin molecules which are present within the tissue. Such crosslinking of the collagen and elastin serves to preserve the tissue so that the tissue may be used as, or incorporated into, bioprosthetic devices intended for long term implantation or attachment to a patient's body. Examples of biological materials which have heretofore been utilized as bioprostheses include cardiac valves, blood vessels, skin, dura mater, pericardium, ligaments and tendons. These anatomical structures typically contain connective tissue matrices, formed of collagen and elastin, and the cellular parenchyma of each tissue is disposed within and supported by its connective tissue matrix.
Each collagen molecule consists of three (3) polypeptide chains which are intertwined in a coiled helical conformation. Chemical fixatives (i.e., tanning agents) used to preserve biological tissues form chemical crosslinkages between functional groups on the polypeptide chains within a given collagen molecules, or between functional groups on adjacent collagen molecules.
When chemical crosslinkages are formed between polypeptide chains within a single collagen or elastin molecule, such crosslinkages are termed "intramolecular", while crosslinkages formed between polypeptide chains of different collagen or elastin molecules are termed "intermolecular".
Elastin fibers are built by crosslinking (natural linkage) of repeating units of smaller molecules in essentially fibrous strands maintained by rigid crosslinking involving desmosine and isodesmosine. Those chemical fixatives which are used to form crosslinkages between the amino groups of collagen molecules also tend to form such crosslinkages between amino groups of elastin molecules. However, the amount of elastin present in most biological tissues is substantially less than the amount of collagen present therein.
Chemical fixative agents which have previously been utilized to crosslink collagen and/or elastin in biological tissues include; formaldehyde, glutaraldehyde, dialdehyde starch, hexamethylene diisocyanate and certain polyepoxy compounds.
Glutaraldehyde is the most widely used agent for fixing biological tissues to be as bioprostheses and there are currently a number of commercially available glutaraldehyde-fixed bioprosthetic devices, such as, heart valves of porcine origin having support frames or stents (Carpentier-Edwards.RTM. Stented Porcine Bioprosthesis; Baxter Healthcare Corporation; Edwards CVS Division, Irvine, Calif. 92714-5686), prosthetic heart valves formed of a metal frame having leaflets formed of bovine pericardial tissue mounted on the frame (e.g., Carpentier-Edwards.RTM. Pericardial Bioprosthesis, Baxter Healthcare Corporation, Edwards CVS Division; Irvine, Calif. 92714-5686) and stentless porcine aortic prostheses (e.g., Edwards.RTM. PRIMA.TM. Stentless Aortic Bioprosthesis, Baxter Edwards AG, Spierstrasse 5, GH6048, Horn, Switzerland).
ii. Calcification of Biological Tissue Grafts & Prostheses
One problem associated with the implantation of bioprosthetic grafts is that they tend to undergo in situ calcification following implantation. Such calcification can result in undesirable stiffening, degradation and premature failure of the bioprosthesis. Both intrinsic and extrinsic calcification have been known to occur, although the exact mechanism(s) by which such calcification occurs is unknown.
The factors which determine the rate at which chemically-fixed bioprosthetic grafts undergo calcification have not been fully elucidated. However, factors which are thought to influence the rate of calcification include:
a) patient's age; PA1 b) existing metabolic disorders (i.e., hypercalcemia, diabetes, etc.); PA1 c) dietary factors; PA1 d) race; PA1 e) infection; PA1 f) parenteral calcium administration; PA1 g) dehydration; PA1 h) distortion/mechanical factors; PA1 i) inadequate anti-coagulation therapy during initial period following surgical implantation; and PA1 j) host tissue responses. PA1 a) treating the biological material with a chemical crosslinking agent to form crosslinkages between and/or within the connective tissue protein molecules present in the biological material; PA1 b) contacting the biological material with a polyamine compound which has some carboxyl (COOH) functionality (e.g., amino acid peptide or protein); PA1 c) contacting the biological material with a carboxyl activating compound to convert at least some of the free carboxyl (COOH) groups present on i) the connective tissue protein molecules and/or ii) the polyamine (e.g., amino acid, peptide or protein) molecules, into chemical groups which will react with amines (e.g., conversion of the carboxyl (COOH) groups to o-acylisourea groups); and, PA1 d) contacting the biological material with heparin such that the heparin will bind to the amino (NH.sub.2) groups present on the polyamine compound which has been directly bonded to or mechanically linked with the connective tissue proteins.
Glutaraldehyde-fixed bioprosthetic grafts have been observed to calcify sooner than grafts which have been fixed by non-aldehyde fixative agents. Thus, non-aldehyde fixatives, such as polyepoxy compounds, may be useful for manufacturing bioprosthetic graft materials which exhibit improved (i.e., lessened) propensity for calcification. Examples of polyepoxy compounds which may be used to crosslink connective tissue proteins include: ethylene, polyethylene glycol diglycidyl ether (Denacol EX-810, Nagase Chemical, Co., Osaka, Japan) and glycerol polyglycidyl ether (Denacol EX-313, Nagase Chemical, Co., Osaka, Japan).
The prior art has also included numerous reports and publications which purport to describe techniques or process which will mitigate in situ calcification of implanted biological tissues. These publications include; U.S. Pat. No. 4,885,005 (Nashef et al.) entitled Surfactant Treatment of Implantable Biological Tissue To Inhibit Calcification; U.S. Pat. No. 4,648,881 (Carpentier et al.) entitled, "Implantable Biological Tissue and Process For Preparation Thereof"; U.S. Pat. No. 4,976,733 (Girardot) entitled, "Prevention of Prosthesis Calcification"; U.S. Pat. No. 4,120,649 (Schechter) entitled, "Transplants"; U.S. Pat. No. 5,002,256 (Carpentier) entitled, "Calcification Mitigation of Bioprosthetic Implants"; EP 103947A2 (Pollock et al.) entitled, "Method For Inhibiting Mineralization of Natural Tissue During Implantation" and WO84/01879 (Nashef et al.) entitled, "Surfactant Treatment of Implantable Biological Tissue to Inhibit Calcification"; and, in Yi, D., Liu, W., Yang, J., Wang, B., Dong, G., and Tan, H.; Study of Calcification Mechanism and Anticalcification On Cardiac Bioprostheses Pgs. 17-22, Proceedings of Chinese Tissue Valve Conference, Beijing, China, June 1995.
iii. Biocompatability of Tissue Grafts & Prostheses
The overall biocompatability (e.g., antigenicity and immunogenicity) of a fixed biological tissue can significantly affect the severity of post-implantation calcification of that tissue, and may also be a factor in the occurrence of other undesirable post-implantation complications or sequelae, such as platelet activation, thrombogenesis, local inflammation, and/or graft failure. The biocompatability (e.g., antigenicity and/or immunogenicity) of a fixed biological tissue is largely dependant upon the chemical make-up of the tissue (i.e., presence of surface antigens), the type of chemical fixative agent used in fixing the tissue, and the particular methods and conditions used during the fixation (i.e., chemical crosslinking of the connective tissue proteins). Biocompatability-related problems which have known to follow the implantation of a chemically fixed biological grafts and/or prostheses may include: local tissue inflammation, platelet aggregation, host rejection and/or enzymatic degradation of the graft or prosthesis. Like calcification, lack of biocompatability may also result in undesirable post-implantation complications or sequelae.
One method which has been proposed for mitigating the potential for thrombogenesis due to non-biocompatability of a biological graft or prosthesis, is the binding of heparin to the collagen and/or elastin of the implanted biological tissue such that the anti-coagulant properties of the heparin will prevent or deter subsequent thrombogenesis. Heparin is a polysaccharide which consists of alternate residues of L-iduronic acid 2-sulfate and 2-deoxy-2-sulfoaminoglucose 6-sulfate. The anticoagulant properties of heparin are probably due to binding of the heparin molecule to thrombin and antithrombin in plasma in a manner which promotes their subsequent combination. Heparin may also affect lipid metabolism by causing lipoprotein lipase to become bound to cell surfaces.
Heparin may be covalently or ionically bound to collagen or elastin. In the ionic approach, heparin is first ionically bound to protamine to form a heparin-protamine complex. The heparin-protamine complex is then introduced into the collagen or elastin matrix. As a result, heparin is slowly released from the graft into the blood stream. Such slow release of heparin from a lumenal graft (e.g., a tubular vascular graft) may facilitate endothelialization of the lumen of the graft.
Patents and patent applications which have described methods for binding or applying heparin to bioprosthetic materials include: U.S. Pat. No. 4,690,973 Production Process of an Antithrombogenic and Antiadhesive Material for Medical Use (Noishiki Y., Kodaira K., Furuse M., Miyata T., Miyamoto T., and Ito H.) issued Sep. 1, 1987; U.S. Pat. No. 4,704,131 Medical Materials (Noishiki Y., and Miyata T.) issued Nov. 3, 1987; U.S. Pat. No. 4,806,595 Method of Preparing Antithrombogenic Medical Materials (Noishiki Y., Kodaira, K., Furuse, M., and Miyata T.) issued Feb. 21, 1989. Also, the following publications have described methods for binding or applying heparin to bioprosthetic materials: Noishiki Y., Nagaoka S., Kikuchi T., and Mori Y., "Application of Porous Heparinized Polymer to vascular Graft", Trans ASAIO 27:213-218, 1981; Miyata T., Noishiki Y., Matsumae M., and Yamane Y., "A New Method to Give An Antithrombogenicity to Biological materials and Its Successful Application to Vascular Grafts", Trans ASAIO 29:363-368, 1983; Noishiki Y., and Miyata T., "Antiadhesive Collagen Membrane with Heparin Slow Release", The 11th Annual Meeting of the Society for Biomaterials, pp. 99, San Diego, Apr. 25-28, 1985; Noishiki Y., and Miyata T., "Successful Animal Study of Small Caliber Heparin-Protamine-Collagen Vascular Grafts", Trans ASAIO 31:102-106, 1985; Noishiki Y., and Miyata T., "A Simple Method to Heparinize Biological Materials" J Biomed Mater Res 20:337-346, 1986; Noishiki Y., and Miyata T., "Antiadhesive Collagen Membrane with Heparin Slow Release", J Bioactive Biocompatible Polymers 2:325-333, 1987; Satoh S., Niu S., Shirakata S., Oka T., and Noishiki Y., "Development of an Autologous Connective Tissue Tube as a Small Caliber Vascular Substitute", Trans ASAIO 34:655-660, 1988; Miyata T., Furuse M., and Noishiki Y., "Biodegradable Antiadhesive Collagen Membrane with Heparin Slow Release" The 3rd World Biomaterials Congress, PP. 528, Apr. 21-25, Tokyo, 1988.
One problem which can arise when attempting to covalently bind heparin to chemically fixed biological tissues is that the fixation process may use up the majority of the available amino (NH.sub.2) groups on the connective tissue protein molecules (e.g., collagen and elastin), thereby leaving an insufficient number of functional amino groups for subsequent binding to the heparin. Additionally, the few functional amino (NH.sub.2) groups which may remain on the connective tissue proteins may be located or situated so as to be less than optimal for subsequent heparin binding.
In view of the above-explained shortcomings of the prior art, there remains a need for the development of new methodologies which will a) enhance the number and/or availability of functional amino groups on the connective tissue proteins, b) optimize the locations of the available amino groups to facilitate their subsequent bonding to heparin, and c) cause heparin to become bound to the chemically fixed connective tissue proteins in a manner which will result in less calcification and/or enhanced biocompatability and/or decreased thrombogenicity of the implanted graft.